The traditional method of determining the presence or absence of microorganisms in liquids such as water, food products such as juices, or pharmaceuticals has been to filter the fluid through a suitable membrane to trap any microorganisms present in the fluid on the surface of the membrane. The membrane is then placed on a growth media plate such as a Petri Dish filled with agar or other suitable media and then incubated for several days to allow colonies to develop from the captured organisms. The plates are then removed and examined visually so that the number of colonies present can be counted. If the number is high enough further tests can be conducted to determine exactly what organisms are present. (Often the mere presence of organisms is not in and of itself an indication that the fluid is unsafe and further work to identify the specific organisms is required.).
Recently, molecular tests such as DNA arrays, nucleic acid amplification and nucleic acid sequencing have been introduced. Traditional identification is done by using selective media and a variety of biochemical tests.
It is known that the simple enumeration of such organisms can be expedited using an ATP-bioluminescence test, such as is taught by U.S. Pat. No. 5,627,042.
It has also been suggested that one may use PNA probe assays to detect the presence and number of specific microorganisms. See U.S. Pat. No. 5,773,571. One drawback to the use of this system is the presence of spots that do not arise from the presence of microorganisms (false positives). Something, yet undetermined, causes the generation of a signal indicating the presence of a target organism when further investigation reveals that in fact no organism is present in that location. These are probably caused by nonspecific binding of probe molecules to the membranes. This renders the test by itself unreliable, as often the detection of even one target organism is enough to fail the fluid being tested.
Further, none of these tests have been able to detect the presence of low levels of contaminating organisms in large volumes of fluids, and rapidly identify the organisms of concern with the same sample and in relatively short time.
What is needed is a simple process to combine the best of both processes with the same sample to determine the number of total organisms present as well as the number of those that are of concern.
The present invention provides such a process.